Tehran University of Medical Sciences Iranian Journal of Public Health 2251-6085 34 Supple 1 2005 03 15 67 68 M Sabourighannad C McCormick A MacDonald D Rowlands M Harris 2015 10 03 The induction of IFN-β expression is the first stage in the innate anti-viral response. In order to investigate the possible effects of HCV proteins on IFN-β signalling, a baculovirus delivery system was developed to introduce the whole genome of HCV genotype 1b into hepatoma cells. The construct used in this study lacks the 3’UTR which is required for HCV replication, thus enabling us to look at the effects of HCV proteins on the IFN-β signalling pathway without inducing IFN-β expression by virtue of the presence of replicating (double-stranded) viral RNA. To facilitate this analysis the expression of the HCV polyprotein was under the control of a tetracycline–responsive promoter coupled to the HCV 5’UTR. As a comparison, we have also generated a recombinant baculovirus containing the culture adapted sub-genomic replicon (FK5.1) also derived from HCV genotype 1b, and a mutant form thereof containing an inactivating mutation within the NS5B (RdRp) coding sequence (termed GND). We first confirmed that HepG2 cells were able to mount an effective IFN-β response. As expected, the baculovirus carrying the FK5.1 replicon induced the production of IFN-β as judged by the use of an IFN-β-promoter luciferase reporter construct, whereas the GND baculovirus and the full-length 3’UTR deletant failed to induce luciferase expression. We then proceeded to analyse the effect of the HCV polyprotein on exogenous induction of the IFN-β promoter (by transfecting cells with poly I/C). These studies demonstrated that neither the HCV polyprotein nor the non-structural proteins of HCV (expressed from the replicon) had any effect on the dsRNA-mediated induction of IFN-β promoter. Secondly we analysed potential effects on the inhibition of the IFN-β response, using an ISRE-luciferase construct. Again we observed no effect of either the complete polyprotein or the sub-genomic replicon. Lastly, we examined the activation of both IRF-3 and NFκB, two transcription factors induced by dsRNA signalling that are key to the activation of transcription from the IFN-β promoter. Intriguingly, both the wild type or GND-mutant replicon blocked the dsRNA-induced activation of IRF-3 and NFκB. In contrast the full-length 3’UTR deletant had no significant effect on either transcription factor. These results suggest a complex interplay between HCV and the IFN-β system that is dependent both on the context of polyprotein expression (full length compared with sub-genomic replicon) and the primary amino acid sequence (culture adapted compared with infectious clone). https://ijph.tums.ac.ir/index.php/ijph/article/view/2999